Abstract

The success of chemotherapy largely depends on the anticancer immune response triggered by tumor cells that succumb to immunogenic cell death (ICD). One of the hallmarks of ICD is premortem autophagy that facilitates the release of adenosine triphosphate from dying cancer cells and acts as a chemoattractant for dendritic cell precursors. Here, we show that the immune response induced by inoculation of cancer cells undergoing ICD in response to the anthracycline mitoxantrone (MTX) can be improved by a short-term fasting regimen (48 hours of starvation) and that this effect is reversed by systemic administration of the autophagy inhibitor dimethyl α-ketoglutarate. Tumor growth reduction by MTX treatment is known to depend on autophagy induction in cancer cells as well as on an intact immune system. We compared the antitumor effects of MTX on autophagy-competent cancers implanted in wild type (WT) or partially autophagy-deficient (Becn1± or Atg4b-/-) mice. While there was no difference in the tumor growth reducing effects of MTX on tumors evolving in WT, Becn1+/- and Atg4b-/- mice, we observed an increase in the toxicity of MTX on Atg4b-/- mice. These results suggest that autophagy in cancer cells (but less so in host cells) is rate-limiting for therapeutically relevant anticancer immune responses, yet has a major role in blunting the life-threatening toxicity of chemotherapy.

Nutrient deprivation improves tumor vaccination efficacy in an autophagy dependent manner. (A). Schematic outline of the tumor vaccination experiment used in this study. MCA205 fibrosarcoma cells, cultured for 16 hours with mitoxantrone (MTX) to trigger immunogenic cell death, were inoculated into the left flank of 6 weeks-old female C57Bl6/mice. After injection, mice were starved for 48 hours and left untreated or administered (intraperitoneally, i.p.) with the autophagy inhibitor dimethyl-2-oxoglutarate (DMKG). At day 7, mice were rechallenged by injection of live MCA205 cells in the right flank and tumor appearance was monitored over time. (B). Representative analysis of the experiment depicted in (A). Data represent the pool of two different experiments. Statistical significance was calculated by means of the likelihood ratio test. * p < 0.05; *** p < 0.001.

Systemic autophagy deficiency does not impact on the efficacy of anthracycline-based immunogenic chemotherapy. (A-C) Wild type (WT) autophagy competent C57BL/6 mice (A) or autophagy deficient Atg4b−/-(B) and Becn1+/-(C) mice were inoculated subcutaneously (s.c.) with murine fibrosarcoma MCA205 cells. When tumors reached a size of 25 mm2, mice received a single injection (i.p) of mitoxantrone (MTX) or an equivalent volume of PBS and tumor growth was routinely assessed. From left to right: average (± S.E.M) tumor growth curves of treated with PBS or MTX-based chemotherapy; tumor size distribution at day 26 of data; individual growth curves from mice treated with MTX or PBS. Data represent a pool of three different experiments. Statistical analysis of tumor growth curves was performed by Wald test whereas tumor size distribution at defined time points was analyzed by means of an unpaired t test. ** p < 0.01; *** p < 0.001.